IP4 is a newly discovered second messenger that we identified to be critical for T cell development and activation. How IP4 transmits T cell receptor signals to induce thymocyte maturation remains unclear. Its chemical similarity to the membrane lipid PIP3 suggests that it co-regulates PIP3-effectors, including the serine/threonine kinase AKT. In this grant, we propose that AKT is an important effector pathway downstream of IP4 that contributes to the development of mature CD4 or CD8 T cells and regulatory T cells. We hypothesize that IP4 promotes AKT activity through release of active AKT from the membrane and by facilitating AKT binding to Calmodulin, a Ca2+ sensing regulator of protein function. To test this hypothesis, we present three specific aims. In the first, we will characterize AKT binding to IP4 and PIP3 individually and by competitive inhibition analysis. Second, we will determine how IP4 regulates AKT localization, activation and phosphorylation of downstream effectors. Lastly, we will examine the importance of Calmodulin binding on AKT function in vivo and determine its dependence on IP4. We hope that elucidating the mechanism by which IP4 regulates AKT activity will allow a better understanding of how developmental signals precisely and differentially induce AKT activation to promote the maturation of different T cell subsets. Our long term objective is to characterize how IP4 relays signals from T cell surface receptors to direct specific and protective immune responses. PUBLIC HEALTH RELEVANCE: Itpks generate IP4, a novel second messenger that is required for T cell development and activation, and has also been genetically linked to autoimmunity. This proposal seeks to understand how IP4 regulation of the proto-oncogene AKT controls T cell function.